Funds are requested to purchase a Seahorse Bioscience XF96 extracellular flux analyzer platereader. The instrument permits the measurement of two fundamentally important parameters, O2 and pH, for assessing the bioenergetic profile of cells or isolated mitochondria. The device permits sequential time course recording simultaneously in a 96-well platereader format with the ability to inject two different compounds automatically, allowing the determination of the rates of O2 consumption (i.e., the rate of mitochondrial oxidative phosphorylation or OxPhos) and extracellular acidosis (indicative of the rate of lactate production by anaerobic glycolysis). Moreover, the device is capable of measuring O2 consumption in suspensions of isolated mitochondria using very small sample amounts, a method we developed and validated in preliminary experiments using an XF-96 instrument on loan from Seahorse Biosciences. With the design of appropriate protocols, the bioenergetic profile of a variety of cell types and mitochondria from different experimental groups has been demonstrated, and differences in substrate selection, sites of metabolic control, and defects in metabolism can be identified on a high throughput platform. The unique ability of this instrument to accelerate the pace of research on energy metabolism has motivated us to request the device as the cornerstone of a Bioenergetic Profiling Core that will complement cellular and tissue level imaging platforms at Johns Hopkins University and serve a variety of highly experienced investigators that are leaders in several disciplines including cardiovascular biology, cancer biology, and aging. Seven major users, who all lead NIH-funded multi-investigator, multidisciplinary research programs, form the core user group, and several other minor users have already been generating data in trial runs on the demo unit. A broad user base is also expected to be built upon establishing the center, as there are numerous investigators with similar research interests at the University. The technology is proven and has been available since 2006;however, this would be the first such instrument to be installed on the campus as a core resource and made available to the entire Hopkins research community. High throughput functional characterization of the bioenergetic profile of cells is an unmet need that has lagged behind the information explosion generated by genomics, proteomics, and metabolomics. Thus, the only hope of understanding the role of an expanding number of disease-associated changes in proteins involved in energy metabolism will be to replace the tedious and time-consuming classical respiration or metabolite assays with multiplexed time-resolved recordings that are enabled by the Seahorse Bioscience instrument.